Probiotic potential of Enterococcus faecalis strains isolated from meconium

107 bacterial isolates with Gram positive staining and negative catalase activity, presumably assumed as LAB, were isolated from samples of meconium of six donors at Roubaix hospital, in the north of France. All these bacterial isolates were identified by MALDI-TOF mass spectrometry as Enterococcus faecalis. However, only six isolates among which E. faecalis 14, E. faecalis 28, E. faecalis 90, E. faecalis 97 and E. faecalis 101 (obtained from donor 3) and E. faecalis 93 (obtained from donor 5) were active against some Gram negative bacteria (GNB) and Gram positive bacteria (GPB), through production of lactic acid and bacteriocin like inhibitory substances (BLIS). The identification of these isolates was confirmed by 16rDNA sequencing and their genetic relatedness was established by REP-PCR and pulsed field gel electrophoresis (PFGE) methods. Importantly, the aforementioned antagonistic isolates were sensitive to various classes of antibiotics tested, exhibited high scores of coaggregation and hydrophobicity, and were not hemolytic. Taken together, these properties render these strains as potential candidates for probiotic applications

Production of recombinant bacteriocin divercin V41 by high cell density Escherichia coli batch and fed-batch cultures

To increase the yield of heterologous production of the class II bacteriocin DvnRV41 with Escherichia coli Origami (DE3) (pLysS/pCR03), induction of bacteriocin gene expression was optimized by varying the inducer isopropyl β-d-thiogalactopyranoside (IPTG) concentration (0-2mM), and controlled batch and fed-batch cultures were tested on a 2-L scale. A concentration of 0.5mM IPTG was found to be optimal for cell growth and bacteriocin production. Shake flask cultivation of E. coli Origami (DE3) (pLysS/pCR03) gave biomass and bacteriocin yields of 1.54 ± 0.06g cdw/l and 18 ± 1mg DvnRV41/l, respectively. Biomass (2.70 ± 0.06 and 6.8 ± 0.6g cdw/l, respectively) and bacteriocin yields (30 and 74mg DvnRV41 per liter, respectively) were both increased with batch and fed-batch compared to shake flask cultures. Bacteriocin yields reported in this study are among the highest published for other heterologous expression systems in shake flask

Carnobacterium: positive and negative effects in the environment and in foods

The genus Carnobacterium contains nine species, but only C. divergens and C. maltaromaticum are frequently isolated from natural environments and foods. They are tolerant to freezing/thawing and high pressure and able to grow at low temperatures, anaerobically and with increased CO2 concentrations. They metabolize arginine and various carbohydrates, including chitin, and this may improve their survival in the environment. Carnobacterium divergens and C. maltaromaticum have been extensively studied as protective cultures in order to inhibit growth of Listeria monocytogenes in fish and meat products. Several carnobacterial bacteriocins are known, and parameters that affect their production have been described. Currently, however, no isolates are commercially applied as protective cultures. Carnobacteria can spoil chilled foods, but spoilage activity shows intraspecies and interspecies variation. The responsible spoilage metabolites are not well characterized, but branched alcohols and aldehydes play a partial role. Their production of tyramine in foods is critical for susceptible individuals, but carnobacteria are not otherwise human pathogens. Carnobacterium maltaromaticum can be a fish pathogen, although carnobacteria are also suggested as probiotic cultures for use in aquaculture. Representative genome sequences are not yet available, but would be valuable to answer questions associated with fundamental and applied aspects of this important genus

Bioprospecting antimicrobials from lactiplantibacillus plantarum: Key factors underlying its probiotic action

Lactiplantibacillus plantarum (L. plantarum) is a well‐studied and versatile species of lactobacilli. It is found in several niches, including human mucosal surfaces, and it is largely employed in the food industry and boasts a millenary tradition of safe use, sharing a long‐lasting relationship with humans. L. plantarum is generally recognised as safe and exhibits a strong probiotic character, so that several strains are commercialised as health‐promoting supplements and functional food products. For these reasons, L. plantarum represents a valuable model to gain insight into the nature and mechanisms of antimicrobials as key factors underlying the probiotic action of health‐promoting microbes. Probiotic antimicrobials can inhibit the growth of pathogens in the gut ensuring the intestinal homeostasis and contributing to the host health. Furthermore, they may be attractive alternatives to conventional antibiotics, holding potential in several biomedical applications. The aim of this review is to investigate the most relevant papers published in the last ten years, bioprospecting the antimicrobial activity of characterised probiotic L. plantarum strains. Specifically, it focuses on the different chemical nature, the action spectra and the mechanisms underlying the bioactivity of their antibacterial and antiviral agents. Emerging trends in postbiotics, some in vivo applications of L. plantarum antimicrobials, including strengths and limitations of their therapeutic potential, are addressed and discussed

Partial Purification and Characterization of the Mode of Action of Enterocin S37: A Bacteriocin Produced by Enterococcus faecalis S37 Isolated from Poultry Feces

The aim of this research was to purify and characterize the mode of action of enterocin S37, a bacteriocin produced by Enterococcus faecalis S37, a strain recently isolated from the chicken feces. Enterocin S37 has a molecular weight comprised between 4 and 5 kDa. It remained active after 1 h at 80oC and at pH values ranging from 4.0 to 9.0. Furthermore, cell-free supernatant of Enterococcus faecalis S37 and purified enterocin S37 were active against Gram-positive bacteria including Listeria monocytogenes EGDe, L. innocua F, Enterococcus faecalis JH2-2, and Lactobacillus brevis F145. The purification of enterocin S37 was performed by ammonium sulfate precipitation followed up by hydrophobic-interaction chromatography procedures. Treatment of enterocin S37 with proteinase K, α-chymotrypsin, and papain confirmed its proteinaceous nature, while its treatment with lysozyme and lipase resulted in no alteration of activity. Enterocin S37 is hydrophobic, anti-Listeria and likely acting by depletion of intracellular K+ ions upon action on KATP channels. This study contributed to gain more insights into the mode of action of enterocins

Antimicrobial Properties, Functional Characterisation and Application of Fructobacillus fructosus and Lactiplantibacillus plantarum Isolated from Artisanal Honey

Honey is a valuable reservoir of lactic acid bacteria (LAB) and, particularly, of fructophilic LAB (FLAB), a relatively novel subgroup of LAB whose functional potential for human and food application has yet to be explored. In this study, FLAB and LAB strains have been isolated from honeys of different floral origins and selected for their broad antimicrobial activity against typical foodborne pathogenic bacteria and spoilage filamentous fungi. The best candidates, two strains belonging to the species Lactiplantibacillus plantarum and Fructobacillus fructosus, were submitted to partial characterisation of their cell free supernatants (CFS) in order to identify the secreted metabolites with antimicrobial activity. Besides, these strains were examined to assess some major functional features, including in vitro tolerance to the oro-gastrointestinal conditions, potential cytotoxicity against HT-29 cells, adhesion to human enterocyte-like cells and capability to stimulate macrophages. Moreover, when the tested strains were applied on table grapes artificially contaminated with pathogenic bacteria or filamentous fungi, they showed a good ability to antagonise the growth of undesired microbes, as well as to survive on the fruit surface at a concentration that is recommended to develop a probiotic effect. In conclusion, both LAB and FLAB honey-isolated strains characterised in this work exhibit functional properties that validate their potential use as biocontrol agents and for the design of novel functional foods. We reported antimicrobial activity, cytotoxic evaluation, probiotic properties and direct food application of a F. fructosus strain, improving the knowledge of this species, in particular, and on FLAB, more generally

Polynucleotide phosphorylase is involved in the control of lipopeptide fengycin production in Bacillus subtilis

Bacillus subtilis is a wealth source of lipopeptide molecules such as iturins, surfactins and fengycins or plipastatins endowed with a range of biological activities. These molecules, designated secondary metabolites, are synthesized via non-ribosomal peptides synthesis (NRPS) machinery and are most often subjected to a complex regulation with involvement of several regulatory factors. To gain novel insights on mechanism regulating fengycin production, we investigated the effect of the fascinating polynucleotide phosphorylase (PNPase), as well as the effect of lipopeptide surfactin. Compared to the wild type, the production of fengycin in the mutant strains B. subtilis BBG235 and BBG236 altered for PNPase has not only decreased to about 70 and 40%, respectively, but also hampered its antifungal activity towards the plant pathogen Botrytis cinerea. On the other hand, mutant strains BBG231 (srfAA−) and BBG232 (srfAC−) displayed different levels of fengycin production. BBG231 had registered an important decrease in fengycin production, comparable to that observed for BBG235 or BBG236. This study permitted to establish that the products of pnpA gene (PNPase), and srfAA− (surfactin synthetase) are involved in fengycin production. © 2018, Springer-Verlag GmbH Germany, part of Springer Nature

Production of Bioactive Peptides by Lactobacillus Species: From Gene to Application

To compensate for their amino acid auxotrophy, lactobacilli have developed the ability to hydrolyze proteins present in their environment. This proteolytic activity not only generates the free amino acids needed by the bacteria, but also a large variety of peptides, some of which are endowed with biological activities. These so-called “bioactive peptides” (BAPs) are interesting from a nutrition and healthcare perspective. The use of lactic acid bacteria (LAB) such as lactobacilli is an effective strategy for production and valorization of new BAPs. The proteolytic activity of lactobacilli is exerted in a strain- and species-dependent manner: each species exhibits different proteinase content, leading to a large variety of proteolytic activities. This underlines the high potential of Lactobacillus strains to produce novel hydrolysates and BAPs of major interest. This review aims at discussing the potential of different Lactobacillus species to release BAPs from fermentation media and processes. Strategies used for peptide production are presented. Additionally, we propose a methodology to select the most promising Lactobacillus strains as sources of BAPs. This methodology combines conventional approaches and in silico analyses

BAGEL2: mining for bacteriocins in genomic data

Mining bacterial genomes for bacteriocins is a challenging task due to the substantial structure and sequence diversity, and generally small sizes, of these antimicrobial peptides. Major progress in the research of antimicrobial peptides and the ever-increasing quantities of genomic data, varying from (un)finished genomes to meta-genomic data, led us to develop the significantly improved genome mining software BAGEL2, as a follow-up of our previous BAGEL software. BAGEL2 identifies putative bacteriocins on the basis of conserved domains, physical properties and the presence of biosynthesis, transport and immunity genes in their genomic context. The software supports parameter-free, class-specific mining and has high-throughput capabilities. Besides building an expert validated bacteriocin database, we describe the development of novel Hidden Markov Models (HMMs) and the interpretation of combinations of HMMs via simple decision rules for prediction of bacteriocin (sub-)classes. Furthermore, the genetic context is automatically annotated based on (combinations of) PFAM domains and databases of known context genes. The scoring system was fine-tuned using expert knowledge on data derived from screening all bacterial genomes currently available at the NCBI. BAGEL2 is freely accessible at http://bagel2.molgenrug.nl

Interactions between Kluyveromyces marxianus from cheese origin and the intestinal symbiont Bacteroides thetaiotaomicron: Impressive antioxidative effects

The effects of yeast Kluyveromyces marxianus S-2-05, of cheese origin, were assessed on the intestine anaerobe symbiont Bacteroides thetaiotaomicron ATCC 29741 to unveil any changes in its antioxidant properties. To this end, these microorganisms were grown and incubated either separately, or co-incubated, under anaerobic atmosphere. Afterwards, the microbial cells were recovered and washed, and extracts were prepared using a sterile detergent solution to mimic the intestine detergent content. The extracts prepared from K. marxianus S-2-05 and reference strain K. marxianus MUCL 29917, grown under different conditions, were assessed for their antioxidant properties against superoxide anion and hydrogen peroxide. Extracts from both yeasts showed antioxidative effects, which were particularly important for K. marxianus S-02-5 after anaerobic incubation. Moreover, K. marxianus S-02-5 displayed a high level of activity against the aforementioned reactive oxygen species, enhancing that of B. thetaiotaomicron ATCC 29741, after the co-incubation process. Two-dimensional polyacrylamide gel electrophoresis was used to separate the proteins extracted. Superoxide dismutase, thiol peroxidase, rubrerythrin -intensively produced by B. thetaiotaomicron induced by the yeast-were identified by mass spectrometry. The antioxidative potential evidenced for K. marxianus S-02-5 is another advantage which could justify the utilization of this strain as a probiotic for countering intestinal inflammatory processes. © 2017 Elsevier Lt